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Presence of activatable Shiga toxin genotype (stx2d) in Shiga toxigenic Escherichia coli from livestock sources

机译：源自牲畜的志贺毒素产性大肠杆菌中存在可活化的志贺毒素基因型（stx2d）

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Stx2d is a recently described Shiga toxin whose cytotoxicity is activated 10- to 1,000-fold by the elastase present in mouse or human intestinal mucus. We examined Shiga toxigenic Escherichia coli (STEC) strains isolated from food and livestock sources for the presence of activatable stx(2d). The stx(2) operons of STEC were first analyzed by PCR-restriction fragment length polymorphism (RFLP) analysis and categorized as stx(2), stx(2c) (vha), stx(2c) (vhb), or stx(2d) (EH250). Subsequently, the stx(2c) (vha) and stx(2c) (vhb) operons were screened for the absence of a PstI site in the stx(2a) subunit gene, a restriction site polymorphism which is a predictive indicator for the stx(2d) (activatable) genotype. Twelve STEC isolates carrying putative stx(2d) operons were identified, and nucleotide sequencing was used to confirm the identification of these operons as stx(2d). The complete nucleotide sequences of seven representative stx(2d) operons were determined. Shiga toxin expression in stx(2d) isolates was confirmed by immunoblotting. stx(2d) isolates were induced for the production of bacteriophages carrying stx. Two isolates were able to produce bacteriophages phi1662a and phi1720a carrying the stx(2d) operons. RFLP analysis of bacteriophage genomic DNA revealed that phi1662a and phi1720a were highly related to each other; however, the DNA sequences of these two stx(2d) operons were distinct. The STEC strains carrying these operons were isolated from retail ground beef. Surveillance for STEC strains expressing activatable stx(2d) Shiga toxin among clinical cases may indicate the significance of this toxin subtype to human health.
机译：Stx2d是最近描述的志贺毒素，其细胞毒性被小鼠或人肠粘液中存在的弹性蛋白酶激活10到1,000倍。我们检查了从食物和牲畜来源分离出的志贺毒素性大肠杆菌（STEC）菌株是否存在可激活的stx（2d）。首先通过PCR限制性片段长度多态性（RFLP）分析来分析STEC的stx（2）操纵子，并将其分类为stx（2），stx（2c）（vha），stx（2c）（vhb）或stx（2d） ）（EH250）。随后，筛选了stx（2c）（vha）和stx（2c）（vhb）操纵子，以确定在stx（2a）亚基基因中是否缺少PstI位点，该位点多态性是stx（ 2d）（可激活的）基因型。鉴定了十二个带有推定的stx（2d）操纵子的STEC分离物，并使用核苷酸测序来确认将这些操纵子鉴定为stx（2d）。确定了七个代表性stx（2d）操纵子的完整核苷酸序列。通过免疫印迹证实了stx（2d）分离物中的志贺毒素表达。诱导stx（2d）分离株产生带有stx的噬菌体。两个分离株能够产生携带stx（2d）操纵子的噬菌体phi1662a和phi1720a。对噬菌体基因组DNA的RFLP分析表明，phi1662a和phi1720a彼此高度相关。但是，这两个stx（2d）操纵子的DNA序列是不同的。从零售碎牛肉中分离出带有这些操纵子的STEC菌株。对临床病例中表达可激活的stx（2d）志贺毒素的STEC菌株进行的监测可能表明该毒素亚型对人类健康的重要性。

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